Cancer initiation can occur when a cell undergoes a genetic change that allows it to survive and reproduce in defiance of normal growth constraints. Cancer progression occurs when these aberrant cells invade and colonize in foreign areas that are normally reserved for other cells. Aneuploidy, the loss or gain of a small number of whole chromosomes, is now thought to be a major contribution to initiation and progression of human cancer. In fact, the most common hallmark of all human solid tumors is the presence of aneuploid cells. It is essential that we understand the mechanisms that cells have developed to prevent aneupoidy, which is produced by inaccurate chromosome segregation. Accuracy depends critically on attachment of mitotic chromosomes via their kinetochores to the mitotic spindle. We are just beginning to understand the molecular nature of this interface, and have made considerable progress in identifying a key kinetochore complex that is essential for kinetochore-microtubule (MT) attachment. The Ndc80 complex is comprised of 4 proteins: Ndc80 (Hec1 in humans for Highly Expressed in Cancer), Nuf2, Spc24, and Spc25. This complex not only has roles in kinetochore-MT attachment, but also in the mitotic checkpoint, an important anti-proliferation mechanism cells use to prevent erroneous cell division. Important for cancer research, human cells depleted of Nuf2 and Hec1 arrest in mitosis and subsequently undergo apoptosis. Hec1 is a key kinetochore component that we will continue to study as recent evidence has indicated that RNA interference against Hec1 inhibits tumor growth in vivo, making it an important target for anti-cancer therapies. We propose to further study Hec1 and the Ndc80 complex by (1) delineating the functional regions of Hec1 with emphasis on its roles in the mitotic checkpoint, in apoptotic cell death, and in kinetochore-MT formation and regulation, (2) identifying and characterizing proteins that interact with Hec1, and (3) determining the role of Nuf2 (also a member of the Ndc80 complex) in human kinetochore function. I will initiate these studies under the mentorship of Professor Ted Salmon and transition into an independent laboratory during the second year of the award. The mentored year will allow me to gain additional tools that will complement my skills in biochemistry and cell biology, so that I will be in a strong position to make significant contributions to the fields of mitosis and cancer biology. [unreadable] [unreadable] [unreadable]